Our long term objectives are to investigate the mechanisms of secretion and actions of secretin and cholecystokinin (CCK) to better understand physiology and pathophysiology of the two gut hormones. This proposal is to test two major hypotheses: 1) the release of secretin and cholecystokinin are luminally mediated by releasing factors secreted from the upper small intestinal mucosa and actions of the two hormones on pancreatic exocrine secretion are controlled by pancreatic islet hormones including insulin, pancreatic polypeptide, somatostatin and glucagon. Pancreatic juice or proteases in the upper small intestine suppress the release of both CCK and/or secretin and pancreatic exocrine secretion in interdigestive and/or intestinal phase. Feedback regulation of exocrine pancreas observed in rats, dogs and humans are found to be hormonally mediated. These observations,suggested the presence of trypsin-sensitive releasing factors for the two hormones. The releasing factors are found in rat upper intestinal secretion as trypsin-sensitive peptides. The presence of these factors will be confirmed further in the rat and dog. First, secretin releasing factor (S-RF) will be isolated from the intestinal acid perfusate or extract of rats and dogs and purified to homogeneity. Their primary structure will be determined. Subsequently, CCK-releasing factors (CCK-RF) will be isolated and purified, and their structure will be determined. Both factors will be custom synthesized and their specific radioimmunoassay will be developed. The mechanism of releases of S-RF or CCK-RF and their actions in vivo and in vitro will then be investigated. In conscious rats, immunoneutralization of circulating insulin with a rabbit anti-insulin serum completely suppresses pancreatic exocrine secretion stimulated by a liquid meal or exogenous secretin and CCK-8. It also suppresses the release of secretin and CCK. The effect of insulin on the actions of secretin and CCK will be investigated in rats, dogs and humans as well as animals and patients with diabetes mellitus. The study will be extended to isolated perfused pancreas models of normal and diabetic rats and dogs to elucidate the mechanism of insulin action and the roles of 3 pancreatic inhibitory hormones on the exocrine pancreas. The proposed study will enrich the fundamental knowledge of physiology and pathophysiology of secretin and CCK and the pancreatic exocrine function, and also help us to better manage patients with impaired digestive and absorptive function as well as diabetes.